2.4. Laboratory measurements

Single-point blood samples were taken from the cubital vein and stored at −80°C in the Integrated Research Biobank of the University Medicine Greifswald (56). The exact time of blood sampling was recorded. Participants of SHIP-TREND-0 but not participants of SHIP-START-2 were asked to fast before blood sampling. Fasting time was calculated in both cohorts by quantifying the time difference between the time of blood sampling and the time of the last reported caloric intake.

Processing of the serum samples to measure the cortisol concentrations is described elsewhere (57). Briefly, serum samples were prepared and frozen at −80°C directly after blood sampling. Samples were thawed and processed before the measurements. To quantify the serum cortisol concentrations, an immunoassay with low-cross reactivity was used on the AVIDA Centaur XP System (Siemens Healthcare Diagnostics, Eschborn, Germany). The coefficients of variation observed during the course of the measurements were low for both analyzed cohorts (30, 57).

White blood cell count (WBC) was measured either on the XT2000, XE 5000 or SE9000 analyzers from Sysmex (Sysmex Deutschland GmbH, Norderstedt, Germany) or on the Advia 2120i (Siemens Healthcare Diagnostics, Eschborn, Germany). Glycated hemoglobin (HbA1c) concentrations were quantified by high-performance liquid chromatography (Bio-Rad Diamat, Munich, Germany). Triglycerides were determined enzymatically (Dimension VISTA, Siemens Healthcare Diagnostics GmbH, Eschborn, Germany). High-density lipoprotein cholesterol (HDL-C) was measured enzymatically after the preparation with phosphotungstic acid/MgCl2.

All assays were performed by skilled personnel according to the manufacturer's instructions.